title: New hyperekplexia mutations provide insight into glycine receptor assembly, trafficking, and activation mechanisms.
abstract: Hyperekplexia is a syndrome of readily provoked startle responses, alongside episodic and generalized hypertonia, that presents within the first month of life. Inhibitory glycine receptors are pentameric ligand-gated ion channels with a definitive and clinically well stratified linkage to hyperekplexia. Most hyperekplexia cases are caused by mutations in the α1 subunit of the human glycine receptor (hGlyR) gene (GLRA1). Here we analyzed 68 new unrelated hyperekplexia probands for GLRA1 mutations and identified 19 mutations, of which 9 were novel. Electrophysiological analysis demonstrated that the dominant mutations p.Q226E, p.V280M, and p.R414H induced spontaneous channel activity, indicating that this is a recurring mechanism in hGlyR pathophysiology. p.Q226E, at the top of TM1, most likely induced tonic activation via an enhanced electrostatic attraction to p.R271 at the top of TM2, suggesting a structural mechanism for channel activation. Receptors incorporating p.P230S (which is heterozygous with p.R65W) desensitized much faster than wild type receptors and represent a new TM1 site capable of modulating desensitization. The recessive mutations p.R72C, p.R218W, p.L291P, p.D388A, and p.E375X precluded cell surface expression unless co-expressed with α1 wild type subunits. The recessive p.E375X mutation resulted in subunit truncation upstream of the TM4 domain. Surprisingly, on the basis of three independent assays, we were able to infer that p.E375X truncated subunits are incorporated into functional hGlyRs together with unmutated α1 or α1 plus β subunits. These aberrant receptors exhibit significantly reduced glycine sensitivity. To our knowledge, this is the first suggestion that subunits lacking TM4 domains might be incorporated into functional pentameric ligand-gated ion channel receptors.
authors.affiliation: University of Queensland, Queensland Brain Institute and School of Biomedical Sciences, Queensland 4072, Australia.
authors.forename: Anna
authors.initials: A
authors.lastname: Bode
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Sian-Elin
authors.initials: SE
authors.lastname: Wood
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Jonathan G L
authors.initials: JGL
authors.lastname: Mullins
authors.affiliation: University of Queensland, Queensland Brain Institute and School of Biomedical Sciences, Queensland 4072, Australia.
authors.forename: Angelo
authors.initials: A
authors.lastname: Keramidas
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Thomas D
authors.initials: TD
authors.lastname: Cushion
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom; Wales Epilepsy Research Network, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Rhys H
authors.initials: RH
authors.lastname: Thomas
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom; Wales Epilepsy Research Network, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: William O
authors.initials: WO
authors.lastname: Pickrell
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom; Wales Epilepsy Research Network, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Cheney J G
authors.initials: CJG
authors.lastname: Drew
authors.affiliation: Department of Paediatrics, Division of Child Neurology, Faculty of Medicine, University of Jordan, Amman 11942, Jordan.
authors.forename: Amira
authors.initials: A
authors.lastname: Masri
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals National Health Service Foundation Trust, Manchester Academic Health Sciences Centre, Manchester M13 9WL, United Kingdom; Manchester Centre for Genomic Medicine, Institute of Human Development, Faculty of Medical and Human Sciences, University of Manchester, Manchester Academic Health Sciences Centre, Manchester M13 9WL, United Kingdom.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Jones
authors.affiliation: Royal Manchester Children's Hospital, Central Manchester University Hospitals National Health Service Foundation Trust, Manchester Academic Health Sciences Centre, Manchester M13 9WL, United Kingdom.
authors.forename: Grace
authors.initials: G
authors.lastname: Vassallo
authors.affiliation: Department of Pediatrics, Copenhagen University Hospital, Rigshospitalet, 2100 Copenhagen, Denmark.
authors.forename: Alfred P
authors.initials: AP
authors.lastname: Born
authors.affiliation: Department of Pediatrics, Division of Child Neurology, Faculty of Medicine, Basşkent University, 06990 Ankara, Turkey.
authors.forename: Fusun
authors.initials: F
authors.lastname: Alehan
authors.affiliation: Institute of Pediatric Neurology, Schneider Children's Medical Center of Israel, Petah Tikva 49202, Israel; Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69987, Israel.
authors.forename: Sharon
authors.initials: S
authors.lastname: Aharoni
authors.affiliation: Neurology Department, Affinity Medical Group, Menasha, Wisconsin 54952.
authors.forename: Gerald
authors.initials: G
authors.lastname: Bannasch
authors.affiliation: Department of Neonatology, University Medical Center of the Johannes Gutenberg University Mainz, D-55099 Mainz, Germany.
authors.forename: Marius
authors.initials: M
authors.lastname: Bartsch
authors.affiliation: Kocaeli University Medical Faculty, Department of Pediatrics, Division of Child Neurology, 41380 Kocaeli, Turkey.
authors.forename: Bulent
authors.initials: B
authors.lastname: Kara
authors.affiliation: Division of Human Genetics, National Health Laboratory Service, and School of Pathology, Faculty of Health Sciences, University of the Witwatersrand, 2000 Johannesburg, South Africa.
authors.forename: Amanda
authors.initials: A
authors.lastname: Krause
authors.affiliation: Department of Psychiatry and Clinical Psychology, Saint George Hospital University Medical Center, Balamand University, Faculty of Medicine, Beirut 1100 2807, Lebanon.
authors.forename: Elie G
authors.initials: EG
authors.lastname: Karam
authors.affiliation: Department of Psychiatry and Clinical Psychology, Saint George Hospital University Medical Center, Balamand University, Faculty of Medicine, Beirut 1100 2807, Lebanon.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Matta
authors.affiliation: Royal Children's Hospital Melbourne, Children's Neuroscience Centre, Royal Children's Hospital, Victoria 3052, Australia.
authors.forename: Vivek
authors.initials: V
authors.lastname: Jain
authors.affiliation: Metabolic Unit, Meyer Children's Hospital, Rambam Medical Center, Technion Faculty of Medicine, Haifa 31096, Israel.
authors.forename: Hanna
authors.initials: H
authors.lastname: Mandel
authors.affiliation: Department of Pediatrics and Adolescent Medicine, Medical University of Vienna, 1090 Vienna, Austria.
authors.forename: Michael
authors.initials: M
authors.lastname: Freilinger
authors.affiliation: Department of Genetics, Children's Hospital of Eastern Ontario, Ottawa, Ontario K1H 8L1, Canada.
authors.forename: Gail E
authors.initials: GE
authors.lastname: Graham
authors.affiliation: Yorkshire Regional Genetic Service, Chapel Allerton Hospital, Leeds, West Yorkshire LS9 7TF, United Kingdom.
authors.forename: Emma
authors.initials: E
authors.lastname: Hobson
authors.affiliation: Neonatal Unit, Bradford Royal Infirmary, Bradford, West Yorkshire BD9 6RJ, United Kingdom.
authors.forename: Sue
authors.initials: S
authors.lastname: Chatfield
authors.affiliation: Génétique, Pôle Mère, Enfant, Centre Hospitalier d'Arras, 62 000 Arras, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Vincent-Delorme
authors.affiliation: Clalit Health Services, Shfaram 20200, Israel.
authors.forename: Jubran E
authors.initials: JE
authors.lastname: Rahme
authors.affiliation: Zlotowski Center for Neuroscience, Ben-Gurion University of the Negev, Beer-Sheva 8410501, Israel.
authors.forename: Zaid
authors.initials: Z
authors.lastname: Afawi
authors.affiliation: Epilepsy Research Centre, Melbourne Brain Centre, Austin Health, Heidelberg 3084, Victoria, Australia.
authors.forename: Samuel F
authors.initials: SF
authors.lastname: Berkovic
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom; Wales Epilepsy Research Network, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Owain W
authors.initials: OW
authors.lastname: Howell
authors.affiliation: Institut de Pathologie et de Génétique ASBL/IRSPG, B-6041 Gosselies, Belgium.
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Vanbellinghen
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom; Wales Epilepsy Research Network, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Mark I
authors.initials: MI
authors.lastname: Rees
authors.affiliation: Department of Neurology Research and Molecular Neuroscience, Institute of Life Science, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom; Wales Epilepsy Research Network, College of Medicine, Swansea University Swansea SA2 8PP, United Kingdom.
authors.forename: Seo-Kyung
authors.initials: SK
authors.lastname: Chung
authors.affiliation: University of Queensland, Queensland Brain Institute and School of Biomedical Sciences, Queensland 4072, Australia. Electronic address: j.lynch@uq.edu.au.
authors.forename: Joseph W
authors.initials: JW
authors.lastname: Lynch
mesh_terms: D019943:Amino Acid Substitution; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D008297:Male; D009127:Muscle Rigidity / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018009:Receptors, Glycine / Q000235:genetics / Q000378:metabolism*
chemical_list: C000657604:GLRA1 protein, human; C114561:GLRB protein, human; D018009:Receptors, Glycine
keywords: Chloride Channels; Cys-Loop Receptors; Glycine Receptors; Neurological Diseases; Patch Clamp Electrophysiology